WJV  Vol.1 No.3 , August 2011
Recent Developments in the Nasal Immunization against Anthrax
Abstract: Bacillus anthracis is the causative agent of anthrax, a bacterial infection with a high mortality rate [1-3]. Although anthrax infection can be cutaneous, gastrointestinal or pulmonary, the pulmonary form is the most deadly [2,3]. Thus, the release of Bacillus anthracis spores that can be inhaled represents a potent bioterrorism threat; the capacity of B. anthracis spores to act as a bioterrorism weapon was demonstrated in 2001, with the intentional infection of 22 persons in the U.S.A. [2,4]. Until recently, the available vaccines were developed to confer protection against cutaneous infection; despite this, these vaccines demonstrated experimental efficacy against pulmonary infection in multiple animal models [1,2]. Nevertheless, there are many limitations for these vaccines to be considered successful and effective vaccine, including the intensity of the required vaccination schedule, the administration route and the presence of local adverse effects experienced after vaccination [1,3,5,6]. To develop more efficient vaccines against pulmonary anthrax, intranasal formulations with adjuvant have been studied. These formulations have advantages because they are easy to administer and because they are expected to induce both systemic and respiratory tract mucosal immune responses. Therefore, the main goal of this review is to compare the different experimental adjuvants used with anthrax antigens and the different approaches regarding the vaccination schedule and consecutive boosters.
Cite this paper: nullS. Jesus and O. Borges, "Recent Developments in the Nasal Immunization against Anthrax," World Journal of Vaccines, Vol. 1 No. 3, 2011, pp. 79-91. doi: 10.4236/wjv.2011.13008.

[1]   A. U. Bielinska, K. W. Janczak, J. J. Landers, P. Makidon, L. E. Sower, J. W. Peterson, et al., “Mucosal Immunization with a Novel Nanoemulsion-Based Recombinant Anthrax Protective Antigen Vaccine Protects against Bacillus Anthracis Spore Challenge,” Infection and Immunity, Vol. 75, No. 8, 2007, pp. 4020-4029. doi:10.1128/IAI.00070-07

[2]   B. R. Sloat and Z. Cui, “Nasal Immunization with Anthrax Protective Antigen Protein Adjuvanted with Polyriboinosinic-Polyribocytidylic Acid Induced Strong Mucosal and Systemic Immunities,” Pharmaceutical Research, Vol. 23, No. 6, 2006, pp. 1217-1226. doi:10.1007/s11095-006-0206-9

[3]   B. R. Sloat, D. S. Shaker, U. M. Le and Z. Cui, “Nasal Immunization with the Mixture of PA63, LF, and a PGA Conjugate Induced Strong Antibody Responses against All Three Antigens,” FEMS Immunology and Medical Microbiology, Vol. 52, No. 2, 2008, pp. 169-179. doi:10.1111/j.1574-695X.2007.00347.x

[4]   D. B. Jernigan, P. L. Raghunathan, B. P. Bell, R. Brechner, E. A. Bresnitz, J. C. Butler, et al., “Investigation of Biote- rrorism-Related Anthrax, United States, 2001: Epidemio- logic Findings,” Emerging Infectious Diseases, Vol. 8, No. 10, 2002, pp. 1019-1028.

[5]   Y. P. Gauthier, J. N. Tournier, J. C. Paucod, J. P. Corre, M. Mock, P. L. Goossens, et al., “Efficacy of a Vaccine Based on Protective Antigen and Killed Spores against Experimental Inhalational Anthrax,” Infection and Im- munity, Vol. 77, No. 3, 2009, pp. 1197-1207. doi:10.1128/IAI.01217-08

[6]   H. Sijun and X. Yong, “Helicobacter Pylori Vaccine: Mucosal Adjuvant & Delivery Systems,” Indian Journal of Medical Research, Vol. 130, No. 2, 2009, pp. 115-124.

[7]   A. M. Friedlander and S. F. Little, “Advances in the Development of Next-Generation Anthrax Vaccines,” Vaccine, Vol. 27 Supplement 4, 2009, pp. D28-D32. doi:10.1016/j.vaccine.2009.08.102

[8]   A. K. Pickering, M. Osorio, G. M. Lee, V. K. Grippe, M. Bray and T. J. Merkel, “Cytokine Response to Infection with Bacillus Anthracis Spores,” Infection and Immunity, Vol. 72, No. 11, 2004, pp. 6382-6389. doi:10.1128/IAI.72.11.6382-6389.2004

[9]   B. R. Sloat and Z. Cui, “Nasal Immunization with a Dual Antigen Anthrax Vaccine Induced Strong Mucosal and Systemic Immune Responses against Toxins and Bacilli,” Vaccine, Vol. 24, No. 40-41, 2006, pp. 6405-6413. doi:10.1016/j.vaccine.2006.06.002

[10]   Y. Zhang, J. Qiu, Y. Zhou, F. Farhangfar, J. Hester, A. Y. Lin, et al., “Plasmid-Based Vaccination with Candidate Anthrax Vaccine Antigens Induces Durable Type 1 and Type 2 T-Helper Immune Responses,” Vaccine, Vol. 26, No. 5, 2008, pp. 614-622. doi:10.1016/j.vaccine.2007.11.072

[11]   J. Enkhtuya, K. Kawamoto, Y. Kobayashi, I. Uchida, N. Rana and S. Makino, “Significant Passive Protective Effect against Anthrax by Antibody to Bacillus Anthracis Inactivated Spores That Lack Two Virulence Plasmids,” Microbiology, Vol. 152, No. Part 10, 2006, pp. 3103-3110. doi:10.1099/mic.0.28788-0

[12]   S. R. Crowe, L. L. Ash, R. J. Engler, J. D. Ballard, J. B. Harley, A. D. Farris, et al., “Select Human Anthrax Protective Antigen Epitope-Specific Antibodies Provide Protection from Lethal Toxin Challenge,” Journal of Infectious Diseases, Vol. 202, No. 2, 2010, pp. 251-260. doi:10.1086/653495

[13]   S. F. Little, B. E. Ivins, P. F. Fellows and A. M. Friedlander, “Passive Protection by Polyclonal Antibodies against Bacillus Anthracis Infection in Guinea Pigs,” Infection and Immunity, Vol. 65, No. 12, 1997, pp. 5171-5175.

[14]   S. Welkos, S. Little, A. Friedlander, D. Fritz and P. Fellows, “The Role of Antibodies to Bacillus Anthracis and Anthrax Toxin Components in Inhibiting the Early Stages of Infection by Anthrax Spores,” Microbiology, Vol. 147, No. Part 6, 2001, pp.1677-1685.

[15]   P. N. Boyaka, A. Tafaro, R. Fischer, S. H. Leppla, K. Fujihashi and J. R. McGhee, “Effective Mucosal Im- munity to Anthrax: Neutralizing Antibodies and Th cell Responses Following Nasal Immunization with Protective Antigen,” The Journal of Immunology, Vol. 170, No. 11, 2003, pp. 5636-5643.

[16]   J. A. Young and R. J. Collier, “Anthrax Toxin: Receptor Binding, Internalization, Pore Formation, and Transl- ocation,” Annual Review of Biochemistry, Vol. 76, 2007, pp. 243-265. doi:10.1146/annurev.biochem.75.103004.142728

[17]   R. J. Cybulski, Jr., P. Sanz and A. D. O'Brien, “Anthrax Vaccination Strategies,” Molecular Aspects of Medicine, Vol. 30, No. 6, 2009, pp. 490-502. doi:10.1016/j.mam.2009.08.006

[18]   B. E. Ivins, M. L. Pitt, P. F. Fellows, J. W. Farchaus, G. E. Benner, D. M. Waag, et al., “Comparative Efficacy of Experimental Anthrax Vaccine Candidates against Inhalation Anthrax in Rhesus Macaques,” Vaccine, Vol. 16, No. 11-12, 1998, pp. 1141-1148. doi:10.1016/S0264-410X(98)80112-6

[19]   T. N. Brahmbhatt, B. K. Janes, E. S. Stibitz, S. C. Darnell, P. Sanz, S. B. Rasmussen, et al., “Bacillus Anthracis Exosporium Protein BclA Affects Spore Germination, Interaction with Extracellular Matrix Proteins, and Hydrophobicity,” Infection and Immunity, Vol. 75, No. 11, 2007, pp. 5233-5239. doi:10.1128/IAI.00660-07

[20]   J. Joyce, J. Cook, D. Chabot, R. Hepler, W. Shoop, Q. Xu, et al., “Immunogenicity and Protective Efficacy of Bacillus Anthracis Poly-Gamma-D-Glutamic Acid Capsule Covalently Coupled to a Protein Carrier Using a Novel Triazine-Based Conjugation Strategy,” The Journal of Biological Chemistry, Vol. 281, No. 8, 2006, pp. 4831- 4843. doi:10.1074/jbc.M509432200

[21]   J. G. Wright, C. P. Quinn, S. Shadomy and N. Messonnier, “Use of Anthrax Vaccine in the United States: Recommen- dations of the Advisory Committee on Immunization Pra- ctices (ACIP),” MMWR-Recommendations and Reports, Vol. 59, No. RR-6, 2009, pp. 1-30.

[22]   V. Schijns and J. Brewer, “Immunopotentiators in Modern Vaccines’ (IMV-II) held in Malaga, Spain, May 18-20, 2005,” Vaccine, Vol. 24, No. 26, 2006, pp. 5391-5392. doi:10.1016/j.vaccine.2006.03.053

[23]   N. A. Twenhafel, “Pathology of Inhalational Anthrax Animal Models,” Veterinary Pathology, Vol. 47, No. 5, 2010, pp. 819-830. doi:10.1177/0300985810378112

[24]   W. M. Gwinn, S. M. Kirwan, S. H. Wang, K. A. Ashcraft, N. L. Sparks, C. R. Doil, et al., “Effective Induction of Protective Systemic Immunity with Nasally Administered Vaccines Adjuvanted with IL-1,” Vaccine, Vol. 28, No. 42, 2010, pp. 6901-6914. doi:10.1016/j.vaccine.2010.08.006

[25]   M. Zeng, Q. Xu and M. E. Pichichero, “Protection against Anthrax by Needle-Free Mucosal Immunization with Human Anthrax Vaccine,” Vaccine, Vol. 25, No. 18, 2007, pp. 3588-3594. doi:10.1016/j.vaccine.2007.01.075

[26]   T. Ebensen and C. A. Guzman, “Immune Modulators with Defined Molecular Targets: Cornerstone to Optimize Rational Vaccine Design,” Human Vaccines, Vol. 4, No. 1, 2008, pp. 13-22. doi:10.4161/hv.4.1.5560

[27]   G. J. Gorse, W. Keitel, H. Keyserling, D. N. Taylor, M. Lock, K. Alves, et al., “Immunogenicity and Tolerance of Ascending Doses of a Recombinant Protective Antigen (rPA102) Anthrax Vaccine: A Randomized, Double- Blinded, Controlled, Multicenter Trial,” Vaccine, Vol. 24, No. 33-34, 2006, pp. 5950-5959. doi:10.1016/j.vaccine.2006.05.044

[28]   B. K. Brown, J. Cox, A. Gillis, T. C. VanCott, M. Marovich, M. Milazzo, et al., “Phase I Study of Safety and Immuno- genicity of an Escherichia Coli-Derived Recombinant Pro- ective Antigen (rPA) Vaccine to Prevent Anthrax in Adults,” PLoS One, Vol. 5, No. 11, p. e13849. doi:10.1371/journal.pone.0013849

[29]   J. D. Campbell, K. H. Clement, S. S. Wasserman, S. Donegan, L. Chrisley and K. L. Kotloff, “Safety, Reacto- genicity and Immunogenicity of a Recombinant Protec- tive Antigen Anthrax Vaccine Given to Healthy adults,” Human Vaccines, Vol. 3, No. 5, 2007, pp. 205-211. doi:10.4161/hv.3.5.4459

[30]   D. G. Bouzianas, “Potential Biological Targets of Bacillus Anthracis in Anti-Infective Approaches against the Threat of Bioterrorism,” Expert Review of Anti-Infective Therapy, Vol. 5, No. 4, 2007, pp. 665-684. doi:10.1586/14787210.5.4.665

[31]   F. Brossier, M. Levy and M. Mock, “Anthrax spores Make an Essential Contribution to Vaccine Efficacy,” Infection and Immunity, Vol. 70, No. 2, 2002, pp. 661-664.

[32]   J. Kubler-Kielb, E. Vinogradov, H. Hu, S. H. Leppla, J. B. Robbins and R. Schneerson, “Saccharides Cross-Reactive with Bacillus Anthracis Spore Glycoprotein as an Anthrax Vaccine Component,” Proceedings of the National Academy of Sciences of the United States of America, Vol. 105, No. 25, 2008, pp. 8709-8712. doi:10.1073/pnas.0803897105

[33]   A. M. H. Jan Holmgren, M. Lebens, J.-B. Sun, F. Anjuère and C. Czerkinsky, “Mucosal Adjuvants Based on Cholera Toxin and E. Coli Heat-Labile Enterotoxin,” In: D. O’Hagan and V. E. Schijns, Eds., Immunopotentiators in Modern Vaccines, 1st Edition, Elsevier Academic Press, 2006, pp. 235-252.

[34]   S. K. Datta, M. Sabet, K. P. Nguyen, P. A. Valdez, J. M. Gonzalez-Navajas, S. Islam, et al., “Mucosal Adjuvant Activity of Cholera Toxin Requires Th17 Cells and Protects against Inhalation Anthrax,” Proceedings of the National Academy of Sciences of the United States of America, Vol. 107, No. 23, 2010, pp. 10638-10643. doi:10.1073/pnas.1002348107

[35]   M. Matsumoto and T. Seya, “TLR3: Interferon Induction by Double-Stranded RNA Including Poly(I:C),” Adv- anced Drug Delivery Reviews, Vol. 60, No. 7, 2008, pp. 805-812. doi:10.1016/j.addr.2007.11.005

[36]   O. Borges, A. Cordeiro-da-Silva, J. Tavares, N. Santarem, A. de Sousa, G. Borchard, et al., “Immune Response by Nasal Delivery of Hepatitis B Surface Antigen and Codelivery of a CpG ODN in Alginate Coated Chitosan Nanoparticles,” European Journal of Pharmaceutics and Biopharmaceutics, Vol. 69, No. 2, 2008, pp. 405-416. doi:10.1016/j.ejpb.2008.01.019

[37]   O. Borges, J. Tavares, A. de Sousa, G. Borchard, H. E. Junginger and A. Cordeiro-da-Silva, “Evaluation of the Immune Response Following a Short Oral Vaccination Schedule with Hepatitis B Antigen Encapsulated into Alginate-Coated Chitosan Nanoparticles,” European Journal of Pharmaceutical Sciences, Vol. 32, No. 4-5, 2007, pp. 278-290. doi:10.1016/j.ejps.2007.08.005

[38]   C. L. Cooper, H. L. Davis, M. L. Morris, S. M. Efler, M. A. Adhami, A. M. Krieg, et al., “CPG 7909, an Immunostimulatory TLR9 Agonist Oligodeoxynucleotide, as Adjuvant to Engerix-B HBV Vaccine in Healthy Adults: A Double-Blind Phase I/II Study,” Journal of Clinical Immunology, Vol. 24, No. 6, 2004, pp. 693-701. doi:10.1007/s10875-004-6244-3

[39]   C. Bode, G. Zhao, F. Steinhagen, T. Kinjo and D. M. Klinman, “CpG DNA as a Vaccine Adjuvant,” Expert Review of Vaccines, Vol. 10, No. 4, 2011, pp. 499-511. doi:10.1586/erv.10.174

[40]   J. Vollmer and A. M. Krieg, “Immunotherapeutic Applica- tions of CpG Oligodeoxynucleotide TLR9 Agonists,” Ad- vanced Drug Delivery Reviews, Vol. 61, No. 3, 2009, pp. 195-204. doi:10.1016/j.addr.2008.12.008

[41]   D. M. Klinman, D. Currie, G. Lee, V. Grippe and T. Merkel, “Systemic but Not Mucosal Immunity Induced by AVA Prevents Inhalational Anthrax,” Microbes and Infection, Vol. 9, No. 12-13, 2007, pp. 1478-1483. doi:10.1016/j.micinf.2007.08.002

[42]   D. M. Klinman, H. Xie, S. F. Little, D. Currie and B. E. Ivins, “CpG Oligonucleotides Improve the Protective Immune Response Induced by the Anthrax Vaccination of Rhesus Macaques,” Vaccine, Vol. 22, No. 21-22, 2004, pp. 2881-2886. doi:10.1016/j.vaccine.2003.12.020

[43]   Y. Krishnamachari and A. K. Salem, “Innovative Strategies for Co-Delivering Antigens and CpG Oligonucleotides,” Advanced Drug Delivery Reviews, Vol. 61, No. 3, 2009, pp. 205-217. doi:10.1016/j.addr.2008.12.013

[44]   G. Jiang, S. B. Joshi, L. J. Peek, D. T. Brandau, J. Huang, M. S. Ferriter, et al., “Anthrax Vaccine Powder Formu- lations for Nasal Mucosal Delivery,” Journal of Pharma- ceutical Sciences, Vol. 95, No. 1, 2006, pp. 80-96. doi:10.1002/jps.20484

[45]   J. Huang, J. A. Mikszta, M. S. Ferriter, G. Jiang, N. G. Harvey, B. Dyas, et al., “Intranasal Administration of Dry Powder Anthrax Vaccine Provides Protection against Lethal Aerosol Spore Challenge,” Human Vaccines, Vol. 3, No. 3, 2007, pp. 90-93. doi:10.4161/hv.3.3.4011

[46]   J. R. Baldridge, P. McGowan, J. T. Evans, C. Cluff, S. Mossman, D. Johnson, et al., “Taking a Toll on Human Disease: Toll-Like Receptor 4 Agonists as Vaccine Adjuvants and Monotherapeutic Agents,” Expert Opinion on Biological Therapy, Vol. 4, No. 7, 2004, pp. 1129-1138. doi:10.1517/14712598.4.7.1129

[47]   S. D. Klas, C. R. Petrie, S. J. Warwood, M. S. Williams, C. L. Olds, J. P. Stenz, et al., “A Single Immunization with a Dry Powder Anthrax Vaccine Protects Rabbits against Lethal Aerosol Challenge,” Vaccine, Vol. 26, No. 43, 2008, pp. 5494-5502. doi:10.1016/j.vaccine.2008.07.062

[48]   S. Wimer-Mackin, M. Hinchcliffe, C. R. Petrie, S. J. Warwood, W. T. Tino, M. S. Williams, et al., “An Intranasal Vaccine Targeting Both the Bacillus Anthracis Toxin and Bacterium Provides Protection against Aerosol Spore Challenge in Rabbits,” Vaccine, Vol. 24, No. 18, 2006, pp. 3953-3963. doi:10.1016/j.vaccine.2006.02.024

[49]   J. B. McLachlan, C. P. Shelburne, J. P. Hart, S. V. Pizzo, R. Goyal, R. Brooking-Dixon, et al., “Mast Cell Activators: A New Class of Highly Effective Vaccine Adjuvants,” Nature Medicine, Vol. 14, No. 5, 2008, pp. 536-541. doi:10.1038/nm1757

[50]   S. N. Abraham and A. L. St John, “Mast Cell-Orchestrated Immunity to Pathogens,” Nature Reviews Immunology, Vol. 10, No. 6, 2010, pp. 440-452. doi:10.1038/nri2782

[51]   B. C. Baudner and D. T. O'Hagan, “Bioadhesive Delivery Systems for Mucosal Vaccine Delivery,” Journal of Drug Targeting, Vol. 18, No. 10, pp. 752-770. doi:10.3109/1061186X.2010.529143

[52]   N. Mishra, A. K. Goyal, S. Tiwari, R. Paliwal, S. R. Paliwal, B. Vaidya, et al., “Recent Advances in Mucosal Delivery of Vaccines: Role of Mucoadhesive/Biode- gradable Polymeric Carriers,” Expert Opinion on Thera- peutic Patents, Vol. 20, No. 5, 2010, pp. 661-679.

[53]   A. U. Bielinska, M. Gerber, L. P. Blanco, P. E. Makidon, K. W. Janczak, M. Beer, et al., “Induction of Th17 Cellular Immunity with a Novel Nanoemulsion Adjuvant,” Critical Reviews in Immunology, Vol. 30, No. 2, pp.189-199.

[54]   P. E. Makidon, S. S. Nigavekar, A. U. Bielinska, N. Mank, A. M. Shetty, J. Suman, et al., “Characterization of Stability and Nasal Delivery Systems for Immunization with Nanoemulsion-Based Vaccines,” Journal of Aerosol Medicine and Pulmonary Drug Delivery, Vol. 23, No. 2, 2010, pp. 77-89.

[55]   P. Leone, C. G. Janson, L. Bilaniuk, Z. Wang, F. Sorgi, L. Huang, et al., “Aspartoacylase Gene Transfer to the Mammalian Central Nervous System with Therapeutic Implications for Canavan Disease,” Annals of Neurology, Vol. 48, No. 1, 2000, pp. 27-38. doi:10.1002/1531-8249(200007)48:1<27::AID-ANA6>3.0.CO;2-6

[56]   B. R. Sloat and Z. Cui, “Strong Mucosal and Systemic Immunities Induced by Nasal Immunization with Anthrax Protective Antigen Protein Incorporated in Liposome- Protamine-DNA Particles,” Pharmaceutical Research, Vol. 23, No. 2, 2006, pp. 262-269. doi:10.1007/s11095-005-9078-7

[57]   Z. Cui and L. Huang, “Liposome-Polycation-DNA (LPD) Particle as a Carrier and Adjuvant for Protein-Based Vaccines: Therapeutic Effect against Cervical Cancer,” Cancer Immunology, Immunotherapy, Vol. 54, No. 12, 2005, pp.1180-1190. doi:10.1007/s00262-005-0685-2

[58]   B. R. Sloat, M. A. Sandoval, A. M. Hau, Y. He and Z. Cui, “Strong Antibody Responses Induced by Protein Antigens Conjugated onto the Surface of Lecithin-Based Nanoparticles,” Journal of Controlled Release, Vol. 141, No.1, 2010, pp. 93-100. doi:10.1016/j.jconrel.2009.08.023

[59]   J. Ali, M. Ali, S. Baboota, J. K. Sahani, C. Ramassamy, L. Dao, et al., “Potential of Nanoparticulate Drug Delivery Systems by Intranasal Administration,” Current Pharm- aceutical Design, Vol. 16, No. 14, pp. 1644-1653. doi:10.2174/138161210791164108

[60]   H. C. Flick-Smith, J. E. Eyles, R. Hebdon, E. L. Waters, R. J. Beedham, T. J. Stagg, et al., “Mucosal or Parenteral Administration of Microsphere-Associated Bacillus An- thracis Protective Antigen Protects against Anthrax In- fection in Mice,” Infection and Immunity, Vol. 70, No. 4, 2002, pp. 2022-2028. doi:10.1128/IAI.70.4.2022-2028.2002

[61]   J. M. Huang, H. A. Hong, H. Van Tong, T. H. Hoang, A. Brisson and S. M. Cutting, “Mucosal Delivery of Antigens Using Adsorption to Bacterial Spores,” Vaccine, Vol. 28, No. 4, 2010, pp. 1021-1030. doi:10.1016/j.vaccine.2009.10.127

[62]   T. D. Nandedkar, “Nanovaccines: Recent Developments in Vaccination,” Journal of Biosciences, Vol. 34, No. 6, 2009, pp. 995-1003. doi:10.1007/s12038-009-0114-3

[63]   S. S. Rao, D. Styles, W. Kong, C. Andrews, J. P. Gorres and G. J. Nabel, “A Gene-Based Avian Influenza Vaccine in Poultry,” Poultry Science, Vol. 88, No. 4, 2009, pp. 860-866. doi:10.3382/ps.2008-00360

[64]   C. O. Tacket, M. B. Sztein, S. S. Wasserman, G. Losonsky, K. L. Kotloff, T. L. Wyant, et al., “Phase 2 Clinical Trial of Attenuated Salmonella Enterica Serovar Typhi Oral Live Vector Vaccine CVD 908-htrA in U.S. Volunteers,” Infection and Immunity, Vol. 68, No. 3, 2000, pp. 1196-1201. doi:10.1128/IAI.68.3.1196-1201.2000

[65]   J. E. Galen, M. Chinchilla, M. F. Pasetti, J. Y. Wang, L. Zhao, I. Arciniega-Martinez, et al., “Mucosal Immu- nization with Attenuated Salmonella enterica Serovar Typhi Expressing Protective Antigen of Anthrax Toxin (PA83) Primes Monkeys for Accelerated Serum Antibody Responses to Parenteral PA83 vaccine,” Journal of Infectious Diseases, Vol. 199, No. 3, 2009, pp. 326-335. doi:10.1086/596066

[66]   A. Ludwig, S. Bauer, R. Benz, B. Bergmann and W. Goebel, “Analysis of the SlyA-Controlled Expression, Subcellular Localization and Pore-Forming Activity of a 34 kDa Haemolysin (ClyA) from Escherichia coli K-12,” Molecular Microbiology, Vol. 31, No. 2, 1999, pp. 557-567. doi:10.1046/j.1365-2958.1999.01196.x

[67]   M. Osorio, Y. Wu, S. Singh, T. J. Merkel, S. Bhattacharyya, M. S. Blake, et al., “Anthrax Protective Antigen Delivered by Salmonella Enterica Serovar Typhi Ty21a Protects Mice from a Lethal Anthrax Spore Challenge,” Infection and Immunity, Vol. 77, No. 4, 2009, pp. 1475-1482. doi:10.1128/IAI.00828-08

[68]   L. W. Baillie, A. L. Rodriguez, S. Moore, H. S. Atkins, C. Feng, J. P. Nataro, et al., “Towards a Human Oral Vaccine for Anthrax: The Utility of a Salmonella Typhi Ty21a-Based Prime-Boost Immunization Strategy,” Vaccine, Vol. 26, No. 48, 2008, pp. 6083-6091. doi:10.1016/j.vaccine.2008.09.010

[69]   J. E. Galen, J. Y. Wang, M. Chinchilla, C. Vindurampulle, J. E. Vogel, H. Levy, et al., “A New Generation of Stable, Nonantibiotic, Low-Copy-Number Plasmids Improves Immune Responses to Foreign Antigens in Salmonella Enterica Serovar Typhi Live Vectors,” Infection and Immunity, Vol. 78, No. 1, 2010, pp. 337-347. doi:10.1128/IAI.00916-09

[70]   C. K. Cote, C. A. Rossi, A. S. Kang, P. R. Morrow, J. S. Lee and S. L. Welkos, “The Detection of Protective Antigen (PA) Associated with Spores of Bacillus Anthracis and the Effects of Anti-PA Antibodies on Spore Germination and Macrophage Interactions,” Microbial Pathogenesis, Vol. 38, No. 5-6, 2005, pp. 209-225. doi:10.1016/j.micpath.2005.02.001