J Appl Biomed 11:105-113, 2013 | DOI: 10.2478/v10136-012-0032-9

Construction and evaluation of a novel Bacillus subtilis spores-based enterovirus 71 vaccine

Yin-Guang Cao1,2, Zhi-Hui Li3, Ying-Ying Yue3, Nan-Nan Song3, Li Peng3, Le-Xin Wang4,*, Xinxin Lu1
1 Clinical Laboratory, Beijing Tongren Hospital, Capital Medical University, Beijing, China
2 Liaocheng People' Hospital of Taishan Medical University, Shandong, China
3 Key Laboratory of Rare and Uncommon Diseases, Institute of Basic Medicine, Shandong Academy of Medical Sciences, Shandong Province, China
4 School of Biomedical Sciences, Charles Sturt University, Wagga Wagga, Australia

Enterovirus 71 (EV71) infection and its associated hand-foot-mouth disease is a significant public health problem. The purpose of this study is to develop a novel vaccine to prevent EV71 infection. Bacillus subtilis spores were engineered to express VP1 protein of EV71 with CotB as carrier protein. The recombination was tested in adult mice for the ability to induce immune responses. Mice were inoculated orally and intranasally simultaneously with the spores. The vaccine's efficacy on stimulating immune responses was evaluated by measuring the titer of anti-VP1 IgG and IgA with enzyme-linked-immunosorbent serologic assay (ELISA), and the number of VP1-specific T cells by ELIS-POT. Serum titers of IgG (0.41±0.05 vs 0.20±0.07) and IgA (0.24±0.02 vs 0.11±0.01) in mice immunized with recombinant CotB-VP1 spores were higher than that of mice immunized with nonrecombinant spores 1A771. Splenocytes from the group of mice receiving VP1 spores vaccination contained 1.69±0.52/104 VP1-specific T cells, which was greater than the 0.06±0.06/104 cells from the group of mice receiving nonrecombinant spores vaccination. In conclusion, B. subtilis spores displaying VP1 of EV71 are effective in stimulating cellular immunity and humoral immunity in mice.

Keywords: enterovirus 71; immunity; vaccine; Bacillus subtilis spores

Received: October 19, 2012; Revised: January 2, 2013; Published: July 31, 2013  Show citation

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Cao Y, Li Z, Yue Y, Song N, Peng L, Wang L, Lu X. Construction and evaluation of a novel Bacillus subtilis spores-based enterovirus 71 vaccine. J Appl Biomed. 2013;11(2):105-113. doi: 10.2478/v10136-012-0032-9.
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    References

    1. Barnes AG, Cerovic V, Hobson PS, Klavinskis LS. Bacillus subtilis spores: a novel microparticle adjuvant which can instruct a balanced Th1 and Th2 immune response to specific antigen. Eur J Immunol. 37: 1538-1547, 2007. Go to original source... Go to PubMed...
    2. Berger J. The age of biomedicine: current trends in traditional subjects. J Appl Biomed. 9: 57-61, 2011. Go to original source...
    3. Carrillo C, Wigdorovitz A, Oliveros JC, Zamorano PI, Sadir AM, Gómez N, Salinas J, Escribano JM, Borca MV. Protective Immune response to foot-and-mouth disease virus with VP1 expressed in transgenic plants. J Virol. 72: 1688-1690, 1998. Go to original source... Go to PubMed...
    4. Caruso A, Flamminio G, Folghera S, Peroni L, Foresti I, Balsari A, Turano A. Expression of activation markers on peripheral-blood lymphocytes following oral administration of Bacillus subtilis spores. Int J Immunopharmacol. 15: 87-92, 1993. Go to original source... Go to PubMed...
    5. Ceragioli M, Cangiano G, Esin S, Ghelardi E, Ricca E, Senesi S. Phagocytosis, germination and killing of Bacillus subtilis spores presenting heterologous antigens in human macrophages. Microbiology. 155: 338-346, 2009. Go to original source... Go to PubMed...
    6. Challacombe SJ. Salivary antibodies and systemic tolerance in mice after oral immunization with bacterial antigens. Ann N Y Acad Sci. 409: 177-193, 1983. Go to original source... Go to PubMed...
    7. Chan L, Parashar U, Lye M, Ong FG, Zaki SR, Alexander JP, Ho KK, Han LL, Pallansch MA, Suleiman AB, Jegathesan M, Anderson LJ. Deaths of children during an outbreak of hand, foot, and mouth disease in Sarawak, Malaysia: clinical and pathological characteristics of the disease. Clin Infect Dis. 31: 678-683, 2000. Go to original source... Go to PubMed...
    8. Chen HF, Chang MH, Chiang BL, Jeng ST. Oral immunization of mice using transgenic tomato fruit expressing VP1 protein from enterovirus 71. Vaccine. 24: 2944-2951, 2006. Go to original source... Go to PubMed...
    9. Chen HL, Huang JY, Chu TW, Tsai TC, Hung CM, Lin CC, Liu FC, Wang LC, Chen YJ, Lin MF, Chen CM. Expression of VP1 protein in the milk of transgenic mice: A potential oral vaccine protects against enterovirus 71 infection. Vaccine. 26: 2882-2889, 2008. Go to original source... Go to PubMed...
    10. Chen SC, Chang HL, Yan TR, Cheng YT, Chen KT. An eight-year study of epidemiologic features of enterovirus 71 infection in Taiwan. Am J Trop Med Hyg. 77: 188-191, 2007. Go to original source...
    11. Ciabattini A, Parigi R, Isticato R, Oggioni MR, Pozzi G. Oral priming of mice by recombinant spores of Bacillus subtilis. Vaccine. 22: 4139-4143, 2004. Go to original source... Go to PubMed...
    12. Cutting SM, Hong HA, Baccigalupi L, Ricca E. Oral vaccine delivery by recombinant spore probiotics. Int Rev Immunol. 28: 487-505, 2009. Go to original source... Go to PubMed...
    13. Duc le H, Hong HA, Cutting SM. Germination of the spore in the gastrointestinal tract provides a novel route for heterologous antigen delivery. Vaccine. 21: 4215-4224, 2003. Go to original source... Go to PubMed...
    14. Duc le H, Hong HA, Uyen NQ, Cutting SM. Intracellular fate and immunogenicity of B. subtilis spores. Vaccine. 22: 1873-1885, 2004. Go to original source... Go to PubMed...
    15. Duc le H, Hong HA, Atkins HS, Flick-Smith HC, Durrani Z, Rijpkema S, Titball RW, Cutting SM. Immunization against anthrax using Bacillus subtilis spores expressing the anthrax protective antigen. Vaccine. 25: 346-355, 2007. Go to original source... Go to PubMed...
    16. Fais S, Pallone F, Nava C, Magnani M. Lymphocyte activation by B. subtilis spores. Boll Ist Sieroter Milan. 66: 391-394, 1987. Go to PubMed...
    17. Fakhry S, Sorrentini I, Ricca E, De Felice M, Baccigalupi L. Characterization of spore forming Bacilli isolated from the human gastrointestinal tract. J Appl Microbiol. 105: 2178-2186, 2008. Go to original source... Go to PubMed...
    18. Foo DG, Alonso S, Phoon MC, Ramachandran NP, Chow VT, Poh CL. Identification of neutralizing linear epitopes from the VP1 capsid protein of Enterovirus 71 using synthetic peptides. Virus Res. 125: 61-68, 2007. Go to original source... Go to PubMed...
    19. Hinc K, Isticato R, Dembek M, Karczewska J, Iwanicki A, Peszyńska-Sularz G, De Felice M, Obuchowski M, Ricca E. Expression and display of UreA of Helicobacter acinonychis on the surface of Bacillus subtilis spores. Microb Cell Fact. 9: 2, 2010. Go to original source... Go to PubMed...
    20. Isticato R, Cangiano G, Tran HT, Ciabattini A, Medaglini D, Oggioni MR, De Felice M, Pozzi G, Ricca E. Surface Display of Recombinant Proteins on Bacillus subtilis Spores. J Bacteriol. 183: 6294-6301, 2001. Go to original source... Go to PubMed...
    21. Kim JH, Roh C, Lee CW, Kyung D, Choi SK, Jung HC, Pan JG, Kim BG. Bacterial surface display of GFP(uv) on Bacillus subtilis spores. J Microbiol Biotechnol. 17: 677-680, 2007. Go to PubMed...
    22. Knecht LD, Pasini P, Daunert S. Bacterial spores as platforms for bioanalytical and biomedical applications. Anal Bioanal Chem. 400: 977-989, 2011. Go to original source... Go to PubMed...
    23. Kosaka T, Maeda T, Nakada Y, Yukawa M, Tanaka S. Effect of Bacillus subtilis spore administration on activation of macrophages and natural killer cells in mice. Vet Microbiol. 60: 215-225, 1998. Go to original source... Go to PubMed...
    24. Lai EM, Phadke ND, Kachman MT, Giorno R, Vazquez S, Vazquez JA, Maddock JR, Driks A. Proteomic analysis of the spore coats of Bacillus subtilis and Bacillus anthracis. J Bacteriol. 185: 1443-1454, 2003. Go to original source... Go to PubMed...
    25. Lee MS, Chang LY. Development of enterovirus 71 vaccines. Expert Rev Vaccines. 9: 149-156, 2010. Go to original source... Go to PubMed...
    26. Lee S, Belitsky BR, Brinker JP, Kerstein KO, Brown DW, Clements JD, Keusch GT, Tzipori S, Sonenshein AL, Herrmann JE. Development of a Bacillus subtilis-Based Rotavirus Vaccine. Clin Vaccine Immunol. 17: 1647-1655, 2010. Go to original source... Go to PubMed...
    27. Li L, Hu X, Wu Z, Xiong S, Zhou Z, Wang X, Xu J, Lu F, Yu X. Immunogenicity of self-adjuvanticity oral vaccine candidate based on use of Bacillus subtilis spore displaying Schistosoma japonicum 26 KDa GST protein. Parasitol Res. 105: 1643-1651, 2009. Go to original source... Go to PubMed...
    28. Luiz WB, Cavalcante RC, Paccez JD, Souza RD, Sbrogio-Almeida ME, Ferreira RC, Ferreira LC. Boosting systemic and secreted antibody responses in mice orally immunized with recombinant Bacillus subtilis strains following parenteral priming with a DNA vaccine encoding the enterotoxigenic Escherichia coli (ETEC) CFA/I fimbriae B subunit. Vaccine. 26: 3998-4005, 2008. Go to original source... Go to PubMed...
    29. Mauriello EM, Duc le H, Isticato R, Cangiano G, Hong HA, De Felice M, Ricca E, Cutting SM. Display of heterologous antigens on the Bacillus subtilis spore coat using CotC as a fusion partner. Vaccine. 22: 1177-1187, 2004. Go to original source... Go to PubMed...
    30. Mauriello EM, Cangiano G, Maurano F, Saggese V, De Felice M, Rossi M, Ricca E. Germination-independent induction of cellular immune response by Bacillus subtilis spores displaying the C fragment of the tetanus toxin. Vaccine. 25: 788-793, 2007. Go to original source... Go to PubMed...
    31. Oberste M S, Nix W A, Maher K, Pallansch M A. Improved molecular identification of enteroviruses by RT-PCR and amplicon sequencing. J Clin Virol. 26: 375-377, 2003. Go to original source... Go to PubMed...
    32. Ogra PL, Fishaut M, Gallagher MR. Viral vaccination via the mucosal routes. Rev Infect Dis. 2: 352-369, 1980. Go to original source... Go to PubMed...
    33. Ooi MH, Wong SC, Lewthwaite P, Cardosa MJ, Solomon T. Clinical features, diagnosis, and management of enterovirus 71. Lancet Neurol. 9: 1097-1105, 2010. Go to original source... Go to PubMed...
    34. Potot S, Serra CR, Henriques AO, Schyns G. Display of Recombinant Proteins on Bacillus subtilis Spores, Using a Coat-Associated Enzyme as the Carrier. Appl Environ Microb. 76: 5926-5933, 2010. Go to original source... Go to PubMed...
    35. Robinson K, Chamberlain LM, Schofield KM, Wells JM, Le Page RW. Oral vaccination of mice against tetanus with recombinant Lactococcus lactis. Nat Biotechnol. 15: 653-657, 1997. Go to original source... Go to PubMed...
    36. Shih SR, Li YS, Chiou CC, Suen PC, Lin TY, Chang LY, Huang YC, Tsao KC, Ning HC, Wu TZ, Chan EC. Expression of capsid protein VP1 for use as antigen for the diagnosis of enterovirus 71 infection. J Med Virol. 61: 228-234, 2000. Go to original source... Go to PubMed...
    37. Tian H, Yang QZ, Liang J, Dong SY, Liu ZJ, Wang LX. Clinical features and management outcomes of severe hand, foot, and mouth disease from an inland Chinese community. Med Princ Pract. 21: 355-359, 2012. Go to original source... Go to PubMed...
    38. Uyen NQ, Hong HA, Cutting SM. Enhanced immunisation and expression strategies using bacterial spores as heat-stable vaccine delivery vehicles. Vaccine. 25: 356-365, 2007. Go to original source... Go to PubMed...
    39. Wu CN, Lin YC, Fann C, Liao NS, Shih SR, Ho MS. Protection against lethal enterovirus 71 infection in newborn mice by passive immunization with subunit VP1 vaccines and inactivated virus. Vaccine. 20: 895-904, 2001. Go to original source... Go to PubMed...
    40. Yu CK, Chen CC, Chen CL, Wang JR, Liu CC, Yan JJ, Su IJ. Neutralizing antibody provided protection against enterovirus type 71 lethal challenge in neonatal mice. J Biomed Sci. 7: 523-528, 2000. Go to original source... Go to PubMed...
    41. Yue YY, Li ZH, Li P, Song NN, Zhao YH, Meng H. Prokaryotic expression and initial identification of enterovirus 71 structural protein VP1. Shandong Med J. 51: 22-24, 2011.

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