Ph.D. candidate
Hoa Thi Trong Vo
Center for Bioscience and Biotechnology, University of Science, Vietnam National University Ho Chi Minh City; Faculty of Natural Sciences, Quy Nhon University
Title: Recombinant expression of a mutant α-toxin (HlaH35LH48L) fused to immunostimulatory peptides in Bacillus subtilis and its immunogenicity in mice
Name(s): Hoa Thi Trong Voa,b,c, Uyen Thao Ngoa,b, Phuc Hoang Lama,b, Trinh Thi Ngoc Nguyena,b, Hoang Duc Nguyena,b
a Center for Bioscience and Biotechnology, University of Science, Ho Chi Minh City, Vietnam
b Vietnam National University Ho Chi Minh City, Vietnam
c Faculty of Natural Sciences, Quy Nhon University, Vietnam
Bacillus subtilis is a generally regarded as safe (GRAS) microorganism, which has been developed as a major host for recombinant protein expression and antigen delivery. Staphylococcus aureus is a major human pathogen responsible for a wide range of infections, while the emergence of methicillin-resistant S. aureus (MRSA) strains has increasingly limited the effectiveness of antibiotic treatment. Despite extensive research, no licensed vaccine against S. aureus is currently available due to the pathogen’s complex immune evasion mechanisms. Alpha-toxin (Hla), a key virulence factor of S. aureus, is considered a promising vaccine antigen; however, its native cytotoxicity limits direct application.
In this study, the mutant HlaH35LH48L antigen was expressed in B. subtilis using a combined genome-integrated and plasmid-based expression strategy to enhance recombinant protein production. The hlaH35LH48L gene fused with the immunostimulatory peptide Co1A was integrated into the genome of BsHT800F. In addition, the inducible plasmid pHT2707 carrying hlaH35LH48L fused with the RS09 peptide was introduced to further enhance recombinant protein production, generating BsHT2804/pHT2707 strain. Recombinant strains were confirmed by colony PCR, and secreted protein expression was analyzed by SDS-PAGE and Western blotting. The combined expression system significantly enhanced secretion of HlaH35LH48L compared with strains carrying only a single expression system. Quantitative analysis demonstrated that BsHT2804/pHT2707 produced 2.215 mg HlaH35LH48L per gram of precipitated secreted protein, representing a 15-fold increase over the genome-integrated system alone.
To evaluate preliminary immunogenicity, concentrated secreted proteins were administered intranasally to Swiss mice. Serum analysis by ELISA showed that immunized mice developed significantly higher levels of S. aureus-specific IgG antibodies compared with non-immunized controls after two immunizations (p < 0.05).
To our knowledge, this is the first report describing a combined genome-integrated and inducible plasmid-based secretion system for enhanced production of HlaH35LH48L fused with immunostimulatory peptides in B. subtilis. These findings suggest that recombinant B. subtilis may serve as a promising platform for mucosal vaccine development against S. aureus infection.
Biography of the presenting author
Hoa Thi Trong Vo is a PhD candidate in Biotechnology at the University of Science, Vietnam National University Ho Chi Minh City. She is also a lecturer and researcher at the Faculty of Natural Sciences, Quy Nhon University. From 2019 to 2020, she received training in animal biotechnology at the University of Évora, Portugal, through the Erasmus+ student exchange program. In 2023, she completed professional training in “Shipping Infectious Substances and Biological Specimens” organized by Sandia National Laboratories (USA) in collaboration with VOHUN Vietnam.
Her current work focuses on engineering Bacillus subtilis to secrete antigenic proteins of Staphylococcus aureus and evaluating immune responses in animal models. Her research interests include microbiology and the applications of microbiology in health and food sciences.
Presenting author details
Full name: Hoa Thi Trong Vo
Contact number: +84 964 661 107
Email: vothitronghoa@qnu.edu.vn
Researchgate: Hoa Vo
ResearcherID: NVM-7848-2025
ORCID: orcid.org/0009-0001-9781-5489
Session name/ number:
Category: Poster presentation