Staphylococcal enterotoxin B (SEB) is one of the most potent Staphylococcus aureus exotoxins (SEs). Due to its conserved sequence and stable structure, SEB might be a good candidate antigen for MRSA vaccines. Although cellular immune responses to SEB are well-characterized, much less is known regarding SEB-specific humoral immune responses, particularly regarding detailed epitope mapping. In this study, we utilized a recombinant nontoxic mutant of SEB (rSEB) and an AlPO4 adjuvant to immunize BALB/c mice and confirmed that rSEB can induce a high antibody level and effective immune protection against MRSA infection. Next, the antisera of immunized mice were collected, and linear B cell epitopes within SEB were finely mapped using a series of overlapping synthetic peptides. Three immunodominant B cell epitopes of SEB were screened by ELISA, including a novel epitope, SEB205-222, and two known epitopes, SEB97–114 and SEB247-261. Using truncated peptides, an ELISA was performed with peptide-KLH antisera, and the core sequence of the three immunodominant B cell epitopes were verified as SEB97-112, SEB207-222, and SEB247-257. In vitro, all of the immunodominant epitope-specific antisera (anti-SEB97-112, anti-SEB207-222 and anti-SEB247-257) were observed to inhibit SEB-induced T cell mitogenesis and cytokine production from splenic lymphocytes of BALB/c mice. The homology analysis indicated that SEB97–112 and SEB207-222 were well-conserved among different Staphylococcus aureus strains. The 3D crystal structure of SEB indicated that SEB97–112 was in the loop region inside SEB, whereas SEB207-222 and SEB247-257 were in the β-slice region outside SEB. In summary, the fine-mapping of linear B-cell epitopes of the SEB antigen in this study will be useful to understand anti-SEB immunity against MRSA infection further and will be helpful to optimize MRSA vaccine designs that are based on the SEB antigen.
References
[1]
Otto M (2012) MRSA virulence and spread(2012). Cellular Microbiology 14: 1513–1521. doi: 10.1111/j.1462-5822.2012.01832.x
[2]
Demei Z, Fu W, Fupin H, Xiaofei J, Yuxing N, et al. (2011) China's CHINET surveillance of bacterial resistance In 2010. Chinese Journal of infection and chemotherapy 5: 326–329.
[3]
Nakamura A, Miyake K, Misawa S, KunoY, Horii T, et al. (2012) Association between antimicrobial consumption and clinical isolates of methicillin-resistant Staphylococcus aureus: a 14-year study. J Infect Chemother 18: 90–95. doi: 10.1007/s10156-011-0302-6
[4]
Klevens RM, Morrison MA, Nadle J, Petit S, Gershman K, et al. (2007) Invasive methicillin-resistant Staphylococcus aureus infections in the United States. JAMA 298: 1763–1771. doi: 10.1001/jama.298.15.1763
[5]
Daum RS, Spellberg B (2012) Progress Toward a Staphylococcus aureus Vaccine. Vaccines 54: 560–567. doi: 10.1093/cid/cir828
[6]
Irina VP, Ellen JB, Victor ER (2010) Staphylococcal Enterotoxins. Toxins 2 2177–2197. doi: 10.3390/toxins2082177
[7]
Fluer FS (2012) Staphylococcus enterotoxins, their properties and role as pathogenicity factors. Zh Mikrobiol Epidemiol Immunobiol 2: 99–108.
[8]
Dinges MM, Orwin PM, Schlievert PM (2000) Exotoxins of Staphylococcus aureus. Clin. Microbiol. Rev 13: 16–34. doi: 10.1128/cmr.13.1.16-34.2000
Breuer K, Wittmann M, Bosche B, Kapp A, Werfel T (2000) Severe atopic dermatitis is associated with sensitization to staphylococcal enterotoxin B (SEB). Allergy 55: 551–555. doi: 10.1034/j.1398-9995.2000.00432.x
[12]
Rossi RE, Monasterolo G (2004) Prevalence of serum IgE antibodies to the Staphylococcus aureus enterotoxins (SAE, SEB, SEC, SED, TSST-1) in patients with persistent allergic rhinitis. Int Arch Allergy Immunol 133: 261–266. doi: 10.1159/000076833
[13]
Karauzum H, Chen G, Abndou L, Mahmoudieh M, Boroun AR, et al. (2012) Synthetic human monoclonal antibodies towards staphylococcal enterotoxin B (SEB) protective against toxic shock syndrome. J. Biol. Chem 287: 25203–25215.13. doi: 10.1074/jbc.m112.364075
[14]
Larkin EA, Stiles BG, Ulrich RG (2010) Inhibition of toxic shock by human monoclonal antibodies against staphylococcal enterotoxin B. PLoS One 10: 1–9. doi: 10.1371/journal.pone.0013253
[15]
Tilahun ME, Kwan A, Natarajan K, Quinn M, Tilahun AY, et al. (2011) Chimeric anti-staphylococcal enterotoxin B antibodies and lovastatin act synergistically to provide in vivo protection against lethal doses of SEB. PLoS One 11 1–8: 16. doi: 10.1371/journal.pone.0027203
[16]
Varshney AK, Wang X, Cook E, Dutta K, Scharff MD, et al. (2011) Generation, characterization, and epitope mapping of neutralizing and protective monoclonal antibodies against staphylococcal enterotoxin B-induced lethal shock. J. Biol. Chem 286: 9737–9747. doi: 10.1074/jbc.m110.212407
[17]
Karauzum H, Chen G, Abndou L, Mahmoudieh M, Boroun AR, et al. (2012) Synthetic Human Monoclonal Antibodies toward Staphylococcal Enterotoxin B(SEB) Protective against Toxic Shock Syndrome. The Journal of biological chemistry 30: 25203–25215. doi: 10.1074/jbc.m112.364075
[18]
Balali-Mood M, Moshiri M, Etemad L (2013) Medical aspects of bio-terrorism. Toxicon.69: 131–142. doi: 10.1016/j.toxicon.2013.01.005
[19]
Rusnak JM, Kortepeter M, Ulrich R, Poli M, Boudreau E (2004) Laboratory exposures to staphylococcal enterotoxin B. Emerg. Infect Dis.10: 1544–1549. doi: 10.3201/eid1009.040250
[20]
Birch H (2011) Antibodies could lead to MRSA vaccine (2011) Chemistry World. 8: 21–21.
[21]
Varshney AK, Wang X, Scharff MD, MacIntyre J, Zollner RS, et al. (2013) Staphylococcal Enterotoxin B–Specific Monoclonal Antibody 20B1 Successfully Treats Diverse Staphylococcus aureus Infections. The Journal of Infectious Diseases 208 2058–66. doi: 10.1093/infdis/jit421
[22]
Lindsay CD, Griffiths GD (2013) Addressing bioterrorism concerns: options for investigating the mechanism of action of Staphylococcus aureus enterotoxin B. Hum Exp Toxicol 32: 606–619. doi: 10.1177/0960327112458941
[23]
Krakauer T (2010) Therapeutic Down-Modulators of Staphylococcal Superantigen Induced Inflammation and Toxic Shock. Toxins 2: 1963–1983. doi: 10.3390/toxins2081963
[24]
Urushibata Y, Itoh K, Ohshima M, Seto Y (2010) Generation of Fab fragment-like molecular recognition proteins against staphylococcal enterotoxin B by phage display technology. Clin Vaccine Immunol 17: 1708–1. doi: 10.1128/cvi.00229-10
[25]
Nishi JI, Kanekura S, Takei S, Kitajima I, Nakajima T, et al. (1997) B Cell Epitope Mapping of the Bacterial Superantigen Staphylococcal Enterotoxin B. The Journal of Immunology 158: 247–254.
[26]
Bhatti AR, Micusanb VV (1999) Production and characterization of anti-peptide monoclonal antibodies with specificity for staphylocokal enterotoxins A and B. Journal of Microbiological Methods 35: 143–149. doi: 10.1016/s0167-7012(98)00110-9
[27]
Kum WWS, Chow AW (2001) Inhibition of Staphylococcal Enterotoxin A–Induced Superantigenic and Lethal Activities by a Monoclonal Antibody to Toxic Shock Syndrome Toxin–1. The Journal of Infectious Diseases 183: 1739–48. doi: 10.1086/320732
[28]
Landrum ML, Neumann C, Cook C, Chukwuma U, Ellis MW, et al. (2012) Epidemiology of Staphylococcus aureus Blood and Skin and Soft Tissue Infections in the US Military Health System, 2005-2010. JAMA 308: 50–59. doi: 10.1001/jama.2012.7139
[29]
Inskeep TK, Stahl C, Odle J, Oakes J, Hudson L, et al. (2010) Oral Vaccine Formulations Stimulate Mucosal and Systemic Antibody Responses against Staphylococcal Enterotoxin B in a Piglet Model. Clinical and Vaccine Immunology 17: 1163–1169. doi: 10.1128/cvi.00078-10
[30]
Varshney AK, Wang X, Cook E, Dutta K, Scharff MD, et al. (2011) Generation, characterization, and epitope mapping of neutralizing and protective monoclonal antibodies against staphylococcal enterotoxin B-induced lethal shock. J Biol Chem 286: 9737–9747. doi: 10.1074/jbc.m110.212407
[31]
Irving MB, Pan O, Scott JK (2001) Random-peptide libraries and antigen-fragment libraries for epitope mapping and the development of vaccines and diagnostics. Current Opinion in Chemical Biology 5: 314–324. doi: 10.1016/s1367-5931(00)00208-8
[32]
Rux JJ, Burnett RM (2000) Type-specific epitope locations revealed by X-ray crystallographic study of type 5 hexon. Molecular Therapy 1: 18–30. doi: 10.1006/mthe.1999.0001
[33]
Mayer M, Meyer B (2001) Group epitope mapping by saturation transfer difference NMR to identify se adenovirus gments of a ligand in direct contact with a protein receptor”. Journal of the American Chemical Society 123: 6108–6117. doi: 10.1021/ja0100120
[34]
Komatsu N, Jackson HM, Chan KF, Oveissi S, Cebon J, et al. (2013) Fine-mapping naturally occurring NY-ESO-1 antibody epitopes in melanoma patients' sera using short overlapping peptides and full-length recombinant protein. Molecular Immunology 54: 465–471. doi: 10.1016/j.molimm.2013.01.014
[35]
Arad G, Hillman D, Levy R, Kaempfer R (2001) Superantigen antagonist blocks Th1 cytokine gene induction and lethal shock. J Leukoc Biol 69: 921–927.
[36]
Kappler J, Kotzin B, Herron L, Gelfand E, Bigler R, et al. (1989) V beta-specific stimulation of human T cells by staphylococcal toxins. Science 244: 811–813. doi: 10.1126/science.2524876
[37]
Baum D, Yaron R, Yellin MJ (1998) TNF-alpha, not CD154 (CD40L), plays a major role in SEB-dependent, CD4(+) T cell-induced endothelial cell activation in vitro. Cell Immunol 190: 12–22. doi: 10.1006/cimm.1998.1380
[38]
Kappler JW, Herman A, Clements J, Marrack P (1992) Mutations defining functional regions of the superantigen staphylococcal enterotoxin B. J Exp Med 175: 387–396. doi: 10.1084/jem.175.2.387
[39]
Hurley JM, Shimonkevitz R, Hanagan A, Enney K, Boen E, et al. (1995) Identification of class II major histocompatibility complex and T cell receptor binding sites in the superantigen toxic shock syndrome toxin1. J Exp Med 181: 2229–2235. doi: 10.1084/jem.181.6.2229
