Bacillus subtilis as Vaccine-vectors

Bacillus subtilis is a vaccine production host and delivery vehicle with several advantages, such as low production costs, direct administration, safety to human consumption, and the production of spores with adjuvant effects. Due to these characteristics, Bacillus subtilis engineered to express foreign antigens on spore surfaces or within the vegetative cells can be used for oral or nasal delivery of heterologous antigens and confer protective immunity.

Introduction of Bacillus subtilis

Bacillus subtilis as Vaccine-vectors– Creative Biolabs

Bacillus subtilis (B. subtilis) is a well-known aerobic bacterium that has been categorized as a GRAS organism (generally recognized as safe) by the FDA. B. subtilis spores contain at least 70 individual proteins, of which 5 proteins (SpoIVA, SpoVM, SpoVID, SafA, and CotE) are essential for proper spore coat formation, called morphogenetic proteins. Other three proteins called CotX, CotY, and CotZ have shown to be morphogenes of the crust. B. subtilis spores known to be highly stable and safe are easy to purify making this spore-based display system a potentially powerful tool for surface expression of antigens. So far four spore coat proteins have been used for displaying antigens on B. subtilis spores include CotB, CotC, CotG, and CotX. Furthermore, heterologous antigens can also be expressed in B. subtilis on the surface and inside vegetative cells.

Why Can Bacillus subtilis Be Used as Vaccine-vectors?

Extensive research has shown that spores of B. subtilis are an ideal platform for antigen delivery for oral vaccines. First, protected by multiple layers of cortex proteins, spores can survive in extreme conditions such as high temperature, ultraviolet radiation, enzymatic and chemical damage, and desiccation. In addition, the cortex coat proteins of the B. subtilis spores (e.g., CotB, CotC, and CotG) can be used as an anchor protein to stably display heterologous antigen on the surface of the spore. In addition, B. subtilis spores are non-pathogenic and non-invasive and are therefore currently used as probiotics and food additives for humans and other animals. Furthermore, B. subtilis spores are considered to have adjuvant activity that enhances the delivery of heterologous antigens to the gastrointestinal tract.

The Design for Bacillus subtilis as Delivery Carrier in Creative Biolabs

Antigen displayed on vegetative cells: This is a typical method to address antigen presentation using bacterial species which is accomplished by genetic fusion with surface proteins to display the target antigen. B. subtilis strains used in this pathway are typically generated using constitutive promoters such as the Prrn promoter, which drives expression of the exogenous gene in vegetative cells. The spore derived from this type of strain can be used for immunization and can germinate inside host phagocytic cells allowing an efficient antigen presentation process.

Antigens displayed on spores: This is the most commonly used alternative to vaccination, which is typically achieved by fusing translational level of antigenic sequence to an endogenous host spore surface protein. Displaying antigens on the surface of a spore offers the advantage that this process does not require secretion through the membrane. This method requires that the expression should occur during the sporulation phase of the mother cells, which is achieved by using sporulation-specific promoters such as those of the coat proteins.

Antigens adsorbed on the spore surface: This method relies on the absorptive capacity of the spore surface due to physicochemical interactions. Therefore, the pure antigen is adsorbed on the spores from the wild-type B. subtilis strain and used for immunization. Creative Biolabs has improved this approach by including the assembly and display of target antigen synthesized in the mother cell compartment but outside the spore as it occurs for the endogenous coat proteins. The method is to express the heterologous protein in the mother cell compartment by the spore-forming specific cry1Aa promoter in the late stage of sporulation. Due to the hydrophobicity and high electrostaticity of the spore surface, it subsequently attaches or adsorbs to the surface of the spore, which acts as a non-selective adsorption matrix. The adsorption method can be improved by some mutant strains, such as the cotH gene mutation, which has a strongly altered spore coat with higher absorption capacity than the wild-type strains.

Graphical representation of antigen expression in B. subtilis spores. Spore structure comprises multiple layers of the spore that surround the genome located at the central core. (A) The antigen can be displayed on the crust by genetic fusion with the coat protein. (B) During the late stage of sporulation, the antigen can be assembled and displayed on the mother cell compartment but outside the spore, being adsorbed on the surface of spore.

Fig 2 Graphical representation of antigen expression in B. subtilis spores. Spore structure comprises multiple layers of the spore that surround the genome located at the central core. (A) The antigen can be displayed on the crust by genetic fusion with the coat protein. (B) During the late stage of sporulation, the antigen can be assembled and displayed on the mother cell compartment but outside the spore, being adsorbed on the surface of spore. (Rosales-Mendoza and Angulo. 2015)

B. subtilis offers the potential to serve in the development of new attractive vaccines in terms of safety, needle-free administration, thermostability and costs. Creative Biolabs has developed genetically modified B. subtilis strains and B. subtilis spores as vaccine vectors to deliver foreign antigens that produce efficient and long-term protection against specific pathogens. If you have any needs, we are your best choice.

Reference

  1. Rosales-Mendoza S and Angulo C. (2015). Bacillus subtilis comes of age as a vaccine production host and delivery vehicle. Expert Review of Vaccines. 14(8), 1-14.

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